Abstract
Sexually transmitted diseases (STD) are among the most common infections worldwide. These bacterial infections have spread predominantly in the developing/underdeveloped countries, the most common being syphilis, gonorrhea and those induced by Chlamydia trachomatis, Ureaplasma urealyticum or Mycoplasma spp. Due to extensive usage of antibiotics in the recent past, these bacteria developed resistance to those commonly used for treatment, such resistant strains becoming a public health problem in a number of countries. It is well documented that bacterial STD agents are difficult to detect using standard culture media because these methods require special conditions and adequate nutrients. Antimicrobial susceptibility testing is, therefore, difficult to obtain in such cases. In recent years, genetic tests have been frequently employed in STD diagnosis. The study of genes that induce resistance to antibiotics using DNA isolated from these bacteria may prove to be a viable alternative. Genetic methods enable the DNA extraction from different biological samples, and both the presence of the bacteria and their resistance to one or more antibiotics can be determined from a single DNA sample. By studying the genes that induce antibiotic resistance and the plasmids that transfer such genes, the mechanism that leads to antibiotic resistance can be elucidated.
Keywords
- STD pathogens
- antibiotic resistance
- genes
1. Introduction
Sexually transmitted diseases (STD) are caused by a wide variety of bacteria, viruses or parasites transferred from one human being to another primarily by vaginal, anal or oral sexual contact. Different STD can coexist or be simultaneously transmitted, the presence of any such infection increasing the risk of contracting other STD types. Prolonged, untreated infections with these pathogens can induce anexitis, endometritis, pelvic peritonitis or spontaneous abortions (in women) and urethritis, prostatitis or epididymitis (in men), respectively.
Reasons for STDs’ recurrence in some sexually active women include the anatomy of the female sex, multiple partners or nonusage of condoms [1]. Such infections present an increasingly serious threat to global public health, especially for mothers or babies, causing severe complications such as cancer of the cervix, spontaneous abortions, premature births, low birth weight or infertility [2]. A number of studies concluded that ulcerative and nonulcerative STDs significantly increase HIV transmission, and drug resistance is complicating the fight against HIV [3]. The presence in the genital tract of such microbial pathogens may be due to a poor hygiene or risky behaviors as mentioned above [4].
Microorganisms make use of a wide range of mechanisms to resist to antibiotics and survive their attack. The emergence of antibiotic resistance is a natural biological phenomenon that occurs as a reaction to the use of antibiotics. Antibiotic resistance results from the bacteria ability to resist to antibiotics as they develop and multiply in media with high antibiotic concentration.
The main causes of antibiotic resistance are their abusive usage and the spread of resistant bacterial strains or genes bearing information able to induce resistance. There are multiple mechanisms leading to antibiotic resistance, one involving resistance plasmids carrying genes responsible for resistance to antibiotics [5, 6]. The phenomenon of antibiotic resistance can also develop via mutations or by acquiring resistance genes from other resistant bacteria [7–9].
Resistance to macrolides observed in the STD strains is often associated with mutations in the ribosomal protein genes [10, 11]. It is assumed that mutations in a specific region of the 23S rRNA play a major role in inducing resistance to macrolides as a result of their interaction with the ribosomes [12, 13].
Treatment failure because of bacterial resistance to the major groups of antibiotics as a result of extensive usage became a serious threat to human health. Increased usage of nucleic acid amplification tests (NAATs) in the diagnosis of STD leads to the elimination of culture-based techniques, depleting the number of live strains available for resistance testing. A lack of routine test-of-cure makes it impossible to distinguish treatment failure from reinfection [14]. The cost of health care for patients with resistant infections is higher due to longer duration of illness, additional tests and use of more expensive drugs.
It is obvious that the microorganisms’ antimicrobial resistance is a dynamic phenomenon which points out the need for updated prevalence and susceptibility data from vast geographic areas being available for relevant institutions [4].
2. Most common bacterial STD agents and their resistance to antibiotics
STD infections are induced by a diversity of bacterial pathoges, including
Infection further leads to diseases such as cervicitis, pelvic inflammatory disease, urethritis, proctitis, lymphogranuloma venereum, trachoma or reactive arthritis. Transmission of STD pathoges is largely uncontrolled as over 70% of the women and 50% of the men infected with
Plasmids are small circular self-replicating DNA particles coexisting in the bacterial cells that are capable of exporting genetic information from the bacterial chromosome. They can transfer genes usually not present in the bacterial chromosome. In prokaryotic organisms, plasmids usually code for genes that facilitate survival of the bacteria in the environment. In chlamydia, apart from their participation in glycogen synthesis and encoding proteins responsible for their virulence, their role has yet to be clearly recognized. Resistance plasmids carry genes responsible for antibiotics resistance. Plasmids and bacteriophages can be employed in the diagnostics of chlamydioses. Plasmids are already used for detection of chlamydial infections. Bacteriophages could be used as therapeutic agents, potentially replacing antibiotics to address the increasing problem of antibiotic resistance among chlamydia. The easy transfer of infectious elementary bodies into other cells can prove to be a useful biotechnological tool in the treatment of chlamydioses, particularly in the case of concealed and latent infections, principally at the time of chlamydial resistance to antibiotics used by choice of treatment (tetracyclines) [5]. According to some authors, chlamydial plasmids are not conjugative plasmids, and they do not code antibiotic resistance and do not share the capacity for integration, except for the integrative plasmid revealed in
Chlamydial infections have high recurrence rates among sexually active populations [19]. However, whether this recurrence rate for the disease is due to reinfection or to persistent infection with the same organism has been difficult to determine [20]. Immunity to chlamydial infections is type specific; once the initial infection was resolved, reinfection is believed to result from exposure to chlamydial strains differing in type from the initial infecting strain [21]. In contrast, persistent infections are considered those in which
Chlamydial infections are very likely to recur even when appropriate drug therapy is in place. Most clinical failures occur due to reinfection or relapse as the bacterial phenotype deviates to persistent, non replicating antibiotic-resistant types able to revert to the typical reticulate body phenotype once treatment is complete [22, 23]. Administration of tetracyclines and macrolides, impeding bacterial translation by binding to the 30S or 50S ribosomal subunits, is viewed as the recommended first-line therapeutic regimen for chlamydial infections [24]. Clinical isolates from patients with recurrent
Other frequently diagnosed STD worldwide is gonorrhoea, exhibiting a recent rise in its global incidence.
While not required for individual patient management, laboratory investigations are instrumental in the choice of antimicrobial agents. Due to a high prevalence of fluoroquinolone-resistant
Resistance to all antimicrobial agents has developed in some
Mycoplasmas, the smallest free-living microorganisms, are another important source of STDs. They belong to the class of
In contrast to other bacteria,
As a major cause of nongonococcal urethritis in men, having been directly linked to cervicitis, endometritis, and pelvic inflammatory disease in women,
Because of extreme difficulties in cultivating
Various investigators noted that resistance to tetracyclines of the mycoplasmas’ is on an uphill trend worldwide [33], a Tunisian study reporting resistance rates of 22.7 and 25% among ureaplasmas and
3. Determining the antibiotics resistance of pathogen STD agents
Although resistance to several antibiotics has been reported in vitro, no evidence that
The emergence of drug resistance in many pathogenic bacteria compromised severely the therapeutic utility of macrolides. The occurrence of macrolide resistance is yet undocumented for many bacterial pathogens. The various molecular mechanisms inducing bacterial resistance can be collectively characterized as involving drug efflux, drug inactivation or alterations in the drug target site. The probability of developing resistance depends on the types and quantities of drug to which these organisms are exposed [10]. For instance, shortly after erythromycin was introduced in therapy in the 1950s not only was resistance in bacterial pathogens observed, but erythromycin-resistant strains were found to be cross-resistant to all other macrolides and also to the chemically unrelated lincosamide and streptogramin B drugs [40].
Resistance to macrolides of the
Many strains of
Lateral gene transfer (LGT) is a means allowing
As for
Several genetic mechanisms are employed by pathogenic microbes for producing variants to counter host defenses. Such a high proportion of urogenital tract isolates presenting amino acid substitutions in the polymorphic
Investigating mutations in the 23S rRNA gene of macrolide-resistant isolates of wild-type
Following PCR amplification of the 23S rRNA gene of the eight wild-type resistant isolates (exhibiting MIC values above the tissue concentration of the antibiotic present in the urogenital system), no resistance-associated mutations were found at 2057 (
While certain other studies reported that mutations observed in clinical strains were also found in laboratory strains, the reverse was found not to be true likely because rRNA mutations leading to drug resistance in a clinical pathogen often become apparent only when a drug therapy fails to eradicate that pathogen [10].
The first reported cases of clinically significant
The MIC (antibiotic concentration level where fluorescent antibody staining failed to highlight typical inclusions after incubation in cell culture) and the minimum chlamydicidal concentration allowing no inclusions on passage in an antibiotic-free medium (MCC) of these antimicrobial agents for these isolates were determined after a subsequent passage of the contents of duplicate unstained wells to a fresh monolayer in antibiotic-free medium.
Fosfomycin is a broad-spectrum antibiotic that irreversibly binds to the active site of
Another study [51] made use of antibiotic resistance assays and whole genome sequencing to interrogate the hypothesis that two clinical isolates (IU824 and IU888) have acquired certain antibiotic resistance mechanisms (tetracycline efflux mediated by the presence of one of the 28 genes, ribosomal protection via carriage of one of the 10
Inability to introduce stably maintained DNA and perform targeted gene manipulation are long-standing limitations slowing progresses in understanding many of the components associated with the basic biology and pathogenesis of
Previously expensive, laborious and largely inaccessible, routine identification of tetracycline resistance loci became possible through the high-throughput whole-genome sequencing technology that emerged in recent years and enabled the low-cost interrogation of a large number of
In bacteria with multiple rRNA (
A wide range of plasmids, from the small 4.2 kb plasmid to larger conjugative plasmids promoting their own transfer between strains and carrying determinants of antibiotic resistance, is associated with
The use of molecular assays for gonococcal antimicrobial resistance surveillance is particularly suited to the fluoroquinolones, where mutations occur at defined bases in the quinolone resistance-determining regions (QRDRs) of the
DNA was extracted from an additional 33 first-pass urine specimens collected from patients. A previously published RT-PCR assay differentiating ciprofloxacin susceptible from nonsusceptible
As infection with
The
This was the first Romanian study on STD pathogens demonstrating the presence of genes providing antibiotic resistance in the
Elsewhere, mutations at various genes such as 23S rRNA,
A Chinese study combined antimicrobial susceptibility determinations with molecular-based analysis of AZM-resistant in
A high potential for macrolide resistance to occur by mutations in the 23S rRNAs of the bacteria was predicted. The target site for macrolides is the large (50S) subunit of the bacterial ribosome. Many cases of macrolide resistance in clinical strains can be linked to the alteration of specific nucleotides in 23S rRNA within the large ribosomal subunit [10]. Findings were that single base substitutions in rRNA provide macrolide resistance [61].
About 2000 women were tested for vaginal infections. About 1536 of the 2000 vaginal swab samples cultivated in the study [4] were infected with at least one microorganism, a positivity rate of 76.80%.
In a 65-month study conducted on 373 patients attending the Clinic of Dermatovenereology in Belgrade from January 2007 to May 2012, urethral and cervical swabs were analyzed for the presence of
Among the 37 isolates of
Despite the evidence, physicians routinely favor macrolide antibiotics, for example, erythromycin when treating
Antibiotic resistance determination of
A reliable method to determine antimicrobial susceptibility of
4. Conclusions
Discovery of antibiotic resistant strains is an important step in choosing the appropriate treatment against bacterial STD after the positive detection of the causative agents. A proper treatment can prevent the spreading of these diseases in the general population. Antimicrobial susceptibility testing proves to be useful in cases when bacteria can be isolated.
The antibiotic resistance of
The study of the Chlamydiae biology is complicated by their intracellular development, but knowledge of the recombination mechanism(s) may enable effective genetic manipulations.
Antimicrobial resistance of microorganisms is a quite dynamic phenomenon, highlighting the need for current prevalence and susceptibility data from various different institutions or geographic areas. The ability to use molecular assays for STD antimicrobial resistance surveillance would offer a significant advantage to public health programs and selected patient management.
Drug therapies can result in strains containing mutations that confer the highest resistance becoming prevalent. In contrast, rRNA mutations in laboratory strains are intentionally created in order to evaluate drug interaction mechanisms. The creation of phenotypes with less effective resistance is possible only under controlled laboratory conditions.
The routine use of genomics in clinical settings is becoming a reality. For STDs, understanding the diversity of the circulating strains and their changing over time pattern is a primary requirement. Comparative genomic studies on chlamydia have demonstrated that it undergoes extensive recombination. Genome projects can also help us understand the phenomenon of antibiotic resistance in STD agents, particularly in the cases of
In recent year, genomics was asked to clarify a number of clinical aspects. Although genome analysis of bacterial STD was not a main issue, it can undoubtedly reveal previously inaccessible aspects of pathogen biology, including changes that lead to antibiotic resistance thereof. Genetic methods enable the DNA extraction from different biological samples, and both the presence of the bacteria and its resistance to one or more antibiotics can be determined from a single DNA sample. By studying the genes that induce antibiotic resistance and the plasmids that transfer such genes the mechanism that leads to antibiotic resistance can be elucidated.
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